Bosco, L

Caradonna, F

Chiarelli, R

Librizzi, M

Luparello, C

Roccheri, MC

Segreto, C.

Spencer, J

Archivio istituzionale della ricerca - Università di Palermo

2i;.a:oZ BO E,i: r .k 6 }.eJri C;;:Uet il,:;iSi aioinf*n scs'F '- 'u' -É § i': iCn'Edase;E'E€Gì;ec>*l-{uiAriri:'rn:;_ .x,{j § ""*"'"=.9É § § *r*t*t*xp*r*rzz** § *ffi s * S.tnf,a;r:i,.r*trori ''').r s LSr:prratcfndrrlrer,;;rni # ; itrS * 3 S At,iiffiGrEvatuatim**,*rrali=iJ:ìiir-'i,iii,''É* E g€,) in Z .g (l) gPo;vvrn*lc,rrsr,d:rluF;ìrroprir .Ef * :'* àk -+ EllE :: F= * ri-J ii C, h A = _ __u: ? §.P8 * §tz =5: * E-IJ l=.= r ù-": :§jÉ *--eÉ, *, *-,* etr L- ,-(J€ > \EÈtr t.. "#( # # ' i g)§&r .'l !"{ ilE§ ft §EB i,'J;{r,r,,,,:* ^Ei:- - à{gpaioeelluiartarc nomaF \ -- :È +.f E'A =-1§'E oEi i Autophasv E ru;q*r*r*rà i Tr"r";'Ij=-" E r-A?G; IÉ€*.i'*,;**E ='l'"{daci É5ÉT'IIJ g F É* ffira+,t.na Expess[ur PryfiF È A- E §* I ÉÉ P frrorehoPa*ris § ;od ou*ru,a::ry= § lii É-;:::::::, H ,$& -{4 q- :**tr X e §Xg §seaUrcl*noeveuPmeirt €,§ ug§ § fil]a§ifid;irs.Frote;ltrE"o§'*.3*r r? ??g. "rE ?5iuli.'o' q::: r*i.-: :1ìTrME§TO Di Seltfiza r ltatr0t"cGi:! : -0S tlr CtiiI4lClÉ a fAiL,Àarutjt||r {yIB:CEF)Congresso"Ricerca di base, interd isciplinare e traslazionale in a mbito Biologico e Biotecnologico,' - pa lermo, 26 - 27 Giugno ZoI4Gene and Protein Signatures Associated to Treatment of MDA-MB23l Breastcancer cells with JAHA, a novel Histone Deacetylases lnhibitorLibrizzi M.,1- Dqbski J.,2 Dadlez M.,2 Spencer J.,3 Luparello C.1-lDipartimento STEBìCEF, Università di patermo (ttaty)2.lnstitute of Biochemistry ond Biophysics, polish Academy of sciences, warsow (potond)3 Deportment of Chemistry, Schoot of Life Sciences, University of Sussex (tJnited Kingdom)*mariangela.librizzi@unipa.it *claudio.luparello@unipa.itKeywords: HDACi, Differential Display PCR, Real Time PCR, Proteomic analysis, gene and protein expression patternsJay Amin hydroxamic acid (JAHA) [1] is a novel metal-based SAHA analogue synthesized in vitro, that showssignificant cytotoxic activity [2] on MDA-M8231 breast cancer cells" To identify protein signatures associatedto its cytotoxic activity, we utilized a proteomic approach to reveal protein expression changes after 1,g, 24and 48 h of exposure. The protein identification was performed by mass spectrometry, and a total of elevendifferentially-expressed proteins were visualized. Subsequently, Differential Display (DD) gene expressionanalysis was used to identify gene signatures in MDA-M8231 human breast cancer cell line after exposure toJAHA. We found two genes, Rad50 (DNA repair protein) and NTRK2 (neurotrophic tyrosine kinase, receptor,type 2), upregulated in treated cell preparations, and five genes that encode for protein kinase C epsilontype, Protein kinase C iota type, Ergic (ER-Golgi intermediated compartiment 2KDa protein), MED25(Mediator of RNA Polymerase ll transcription subunit25) and Brefeldine A-inhibited guanine nucleotide-exchange protein 3 that were significantly down-regulated after treatment with JAHA.The result obtained byDD-PCR will be confirmed by Real Time PCR analysis. Further study will be required to compare the reportedsignature pattern with that obtained after exposure of MDA-M8231 cells with the parental molecule SAHA,and to understand the biological implications of the expression changes found.[1] Spencer et al. (2011) . ACS Med Chem Lett. Z,35g-62.l2l Librizzi et a l. (20 1 2 ) . Ch e m. R e s. Toxi c o l. ZS, 2608-16.The effect of the HDAci JAHA on DNA Methytation of breast cancer celts by down-regulating DNMTl through ERK signalingLibrizzi M.,l Chiarelli R", 1 SegrelLo C.,f* Caradonna F., 1 Spencer J.,2 Bosco 1.,1 Roccheri M.C.,l Luparello C.1-lDipartimento STEBiCEF, L)niversità di Polermo lttalyl2Oepoirtment oj Chemistry, Schoot of Life Sciences, lJniversity ofsussex (united Kingdom) *claudiasegreto@libero.it, *claudio.luparello@unipa.itKeywords: HDACi, DNA Methylation, DNMTl level, ERK signaling, MDA-M8231 breast cancer cellsMethylation of CpG repeats in the upstream/promoter regions of genes is an established mechanism of genesilencing in many cell types" DNA methylation results in the recruitment of histone deacetylases (HDACs) topromoter regions, thereby repressing expression of genes. General inhibitors of class I and ll HDACs (HDAC|),suppress the growth of cancer cells in vitro and in vivo. ln this study, we investigated the effect of JAHA [1],a novel HDACi, on the intracellular signaling pathways of MDA-M8231 breast cancer cells" Concerning theMEK pathwayJAHA repressed MAP kinase (ERK) activation after 18 h until 30 h of the treatment, and alsodown-regulated DNA (cytosine-5-)-methyltransferase 1 (DNMT1), a downstream ERK target, already at 1ghwith an increase up to 48 h. To check the occurrence of changes in the extent of global DNA methylation,genomic DNA was submitted to MeSAP (Methylation Sensitive Restriction Arbitrarily-primed) pCR [2] usingAfa and then Hpall enzymes followed by PCR amplification with an arbitrary primer binding preferentially toguanine and cytysine (GC)-rich regions of DNA, including CpG islands" preliminary indications suggest theability of JAHA to induce hypomethylation patterns in tumoral breast cancer cells after 30 h of thetreatment. Collectively, these data demonstrate that the HDACI JAHA, by inhibiting ERK activity, regulatesDNMTl expression and ultimately DNA methylation"[1]Spencer J. et al. (2011). ACS Med Chem 1ett.2,358-62.[2] Naselli F. et al" (2OL4).Gene 535: 29-39Poster Presentation 35

The effect of the HDACi JAHA on DNA methylation of breast cancer cells by down-regulating DNMT1 through ERK signaling

https://iris.unipa.it/retrieve/handle/10447/96010/120816/congresso%20IBIM-STEBICEF%202014-2.pdf

The effect of the HDACi JAHA on DNA methylation of breast cancer cells by down-regulating DNMT1 through ERK signaling

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Archivio istituzionale della ricerca - Università di Palermo